CD5+/Mac-1- peritoneal B cells: a novel B cell subset that exhibits characteristics of B-1 cells

The peritoneal cavity of mice is enriched for B-1 B cells, a lymphocyte subset that differs from conventional B-2 cells phenotypically, functionally, and developmentally. According to current paradigms, all peritoneal B-1 cells express Mac-1 whereas B-2 cells do not and thus these populations are often purified by FACS sorting or magnetic bead isolation based on B cell expression of Mac-1 or lack thereof. However, in the course of studying B220+/Mac-1- peritoneal B-2 cells, we discovered that this population is actually heterogeneous, with approximately 30-40% of these B220+/Mac-1- cells expressing the B-1 cell marker CD5. It was unclear whether this B220+/CD5+/Mac-1- peritoneal B cell population represented aberrantly CD5 expressing B-2 cells or Mac-1- B-1 cells. To address this issue we tested CD5+/Mac-1- peritoneal B cells for several traits that distinguish B-1 and B-2 cells. We found that CD5+/Mac-1- peritoneal B cells resembled CD5+ B-1 cells and not B-2 cells in terms of expression of several additional surface markers (IgM, IgD, CD23, CD43, and CD80). Further, CD5+/Mac-1- peritoneal B cells expressed high levels of V(H)11 and V(H)12, two Ig variable genes that are expressed mainly by B-1 but not B-2 cells. In addition, CD5+/Mac-1- peritoneal B cells responded to PMA, a mitogen that stimulates B-1 cells but not B-2 cells, and not to anti-Ig, that stimulates B-2 cells but not B-1 cells. ELISPOT analyses of freshly isolated CD5+/Mac-1- peritoneal B cells revealed that they secreted IgM constitutively, like B-1 cells and unlike B-2 cells. These results indicate that CD5+/Mac-1- peritoneal B cells are a new subset of B-1 cells, here termed B-1c, and stress the importance of using multiple surface markers to identify and purify specific B cell populations.     

Age-dependent increase of peritoneal B-1b B cells in SCID mice

The impact of increasing age upon immunoglobulin production and B-lymphocyte generation in "leaky" severe combined immune-defective (SCID) mice was examined by enzyme-linked immunosorbent assay and flow cytometry. By 1 year of age, the mice had normal numbers of B cells in their peritoneal cavity, while their spleen had very few immunoglobulin M-positive (IgM+) cells. The majority of B cells expressed the CD11b marker characteristic of the B-1b subset. B-1a (CD5+) cells were present at a lower frequency and B-2 cells were absent. The frequency of mice producing detectable immunoglobulin increased with age, and isotype diversity within individual mice was variable. IgM production was most frequently observed followed by IgG3 and IgG2a, then IgG1, and finally IgA. The selective persistence of the B-1 B-cell subset in the peritoneal cavity of aging SCID mice is a natural model for the study of those genetic and environmental influences that determine lymphocyte longevity.     

X-chromosome-linked immune-deficient mice have B-1b cells

The B lymphocyte subsets of X-chromosome-linked immune-deficient (XID) mice were examined by flow cytometric analyses of spleen and peritoneal cells. As shown in prior studies, young adult XID mice had reduced representation of the CD5+ (B-1a) subset in their peritoneal cavity. However, the CD11b+ (B-1b) B-cell subset was present and exhibited the IgM(hi) CD45(lo) CD23- phenotype characteristic of most B-1 cells. Although present at a lower frequency than that found in their normal counterparts, B-1b cells were evident in CBA/N and (XD2J)F1 male mice. With increasing age, B-1b cell number increased and in the oldest XID mice were present as B-cell chronic lymphocytic leukaemia. These results show that XID mice do have B-1 cells, particularly the B-1b subset.     

B-1 B cell subset composition of DBA/2J mice

Studies of B cell subpopulations have focused upon BALB/c mice and related strains. The B cell subset composition of DBA/2J mice, a prototype strain for BALB/c mice, has been investigated less thoroughly. This report provides the results of a study of the B-1 B cells of DBA/2J mice. In contrast to C.B-17 mice, in which B-1 B cells expressed both the CD5 and CD11b antigens, CD11b expression was most characteristic of DBA/2J B-1 B cells. This was particularly evident in the peritoneal cavity where CD5-CD11b+ B cells were the predominant B cell subpopulation. The number of B-1 B cells increased with age in both the spleen and peritoneal cavity. Strain-specific differences in B cell subset composition may be significant when considering B cell lymphomagenesis with aging.     

B-1 cells in the bone marrow are a significant source of natural IgM

Natural IgM antibodies secreted in the absence of antigenic challenge are important contributors to antimicrobial immunity and tissue homeostasis. Early studies identified BM and, to a lesser extent the spleen, as main tissue sources of this spontaneously secreted IgM. However, the responsible B-cell subset has never been identified. Using multicolor flow cytometry, cell sorting and chimeric mice in which B-1 and B-2 cells and their secreted antibodies are distinguished by their Ig-allotype, we unequivocally identify the natural IgM-secreting cells in spleen and, for the first time, in the BM as IgM(+) IgD(lo/-) CD19(hi) CD43(+) CD5(+/-) B-1 cells. The newly identified population of BM B-1 cells shows many of the phenotypic characteristics of splenic B-1 cells but is distinct from B-1 cells in the peritoneal cavity, which generate at best very small amounts of IgM. Antibody-secreting spleen and BM B-1 cells are distinct also from terminally differentiated plasma cells generated from antigen-induced conventional B cells, as they express high levels of surface IgM and CD19 and lack expression of CD138. Overall, these data identify populations of non-terminally differentiated B-1 cells in spleen and BM as the most significant producers of natural IgM.     

B-1a cells are imprinted by the microenvironment in spleen and peritoneum

B-1a cells are found mainly in the peritoneal cavity of mice but are also present in the spleen. Gene expression profiling defined many genes differentially expressed in B-1a cells from these two sites. To see whether this gene expression pattern was imprinted by the particular microenvironment, peritoneal or spleen cells from recombinant L2 mice mainly consisting of B-1a cells were adoptively transferred into Rag1-/- mice. Re-isolated peritoneal and splenic B-1a cells were analyzed for expression of three indicator genes--vcam-1, adamdec1 and spi-c. The expression of these genes was up-regulated in splenic and down-regulated in peritoneal cells. This particular pattern was observed for peritoneal or splenic donor cells transferred either intraperitoneally or intravenously. Similar results were obtained when levels of surface IgM or frequencies of Mac-1+ B-1 cells were compared after transfer. This suggests that the environment induces the particular genetic program of B-1a cells and argues against an independent ontogeny.     

Up-regulation of CD80-CD86 and IgA on mouse peritoneal B-1 cells by porin of Shigella dysenteriae is Toll-like receptors 2 and 6 dependent

Porin of Shigella dysenteriae type 1 increased the mRNA levels for Toll-like receptor (TLR) 2 and TLR6 by 1.5- and 2.9-fold respectively, of peritoneal cavity B-1a and B-1b cells, implicating that coexpression of TLR2 and TLR6 is essential as a combinatorial repertoire for recognition of porin by the B-1 cells. Among the two key TLRs, TLR2 and TLR4, which are primarily responsible for recognizing majority of the bacterial products, TLR2 and not TLR4, participates in porin recognition. TLR2 got increased on both the B-1 cell populations whereas the TLR4 expression remained unaffected. Besides TLRs, mRNA for MyD88, an effector molecule associated with TLR-mediated response was enhanced by 1.8-fold that suggests of its involvement in the activity of porin. Both of the B-1 cell populations expressed strongly the mRNA for NF-κB in the presence of porin, that was 2.4-fold more than untreated control, conforming to the earlier finding that coexpression of TLR2 and TLR6, resulted in robust NF-κB activation for signaling. Porin treatment of B-1 cell populations of C57BL/6 mice, and C3H/HeJ mice in particular, selectively up-regulated the expression of the costimulatory molecules. CD80 expression got enhanced on the B-1a cells whereas CD86 got solely expressed on B-1b cells. Porin-induced cell surface expression of IgM and IgA on B-1 cell populations from C57BL/6 mice. The IgA-generating capacity, hallmark of mucosal immune response, was confirmed with B-1 cells of C3H/HeJ, the lipopolysaccharide non-responder mouse, in response to the protein. The porin-mediated induction of IgA was augmented by interleukin-6 on B-1a and B-1b cells, by 2.4- and 2.6-fold, respectively. The IgA expressed on both B-1a and B-1b cell surfaces after 72 h of culture was found to bind to the 38 kDa monomer of porin confirming it to be anti-porin IgA antibody.     

LPS-induced migration of peritoneal B-1 cells is associated with upregulation of CXCR4 and increased migratory sensitivity to CXCL12

B-1 cells, which constitute a predominant lymphocyte subset in serosal cavities and produce most of natural antibodies, are subdivided into the CD5(+) B-1a and CD5(-) B-1b cell subpopulations, but the differential roles of B-1a and B-1b cells are not well understood. We report that B-1a cells preferentially migrate out of the peritoneal cavity and upregulate the expression of CXCR4 with heightened sensitivity to CXCL12 and CXCL13 upon LPS treatment compared to B-1b and B-2 cells. Whereas B-1a cells were slightly more abundant than B-1b and B-2 cells in the homeostatic condition, the number of B-1a cells preferentially decreased 48 hr after LPS treatment. The decrease in the peritoneal B-1a cell number was accompanied with increased migration of B-1a cells toward CXCL-12 and CXCL-13 in in vitro transmigration assay using peritoneal B cells from LPS treated mice. The expression level of CXCR4, but not of CXCR5, was also more prominently increased in B-1a cells upon LPS stimulation. LPS-stimulated B-1a cells did not accumulate in omental milky spots in contrast to B-2 cells. These results suggest that B-1a cells actively migrate out of the peritoneal cavity through the regulation of the migratory responsiveness to chemokines and actively participate in systemic immune responses.     

Reduced susceptibility to Fas-mediated apoptosis in B-1 cells

Elimination of activated T and B cells by Fas-dependent apoptosis may contribute to the maintenance of peripheral tolerance. CD40 ligation was recently shown to up-regulate Fas expression and enhance susceptibility to Fas-mediated apoptosis in mouse splenic B cells. In the present study, we have investigated the regulation of Fas expression and Fas-triggered apoptotis in mouse peritoneal B-1 cells. B-1 cells expressed a similar level of CD40 as that on B-2 cells, and proliferated in response to a soluble CD40 ligand (CD40L)-CD8α chimeric protein, suggesting that CD40 on B-1 cells is functional. In contrast to B-2 cells, B-1 cells expressed Fas at only low levels in response to CD40L-CD8α alone or CD40L-CD8α+interleukin-4, and were resistant to Fas-mediated apoptosis following these treatments. While Fas expression could be induced in B-1 cells to a comparable level as that in B-2 cells by cross-linking CD40L-CD8α with an anti-CD8α antibody, the sensitivity to Fas-mediated apoptosis in B-1 cells was significantly reduced compared with B2 cells. These results suggest that peritoneal B-1 cells from normal mice have a lower susceptibility to Fas-mediated apoptosis and may distinguish B-1 from B-2 cells. Similarly, B-1 cells from the peritoneal cavity and spleen of autoimmune-prone NZB mice exhibited reduced susceptibility to Fas-mediated apoptosis relative to their B-2 counterparts. NZB splenic B-1 cells, however, were more susceptible to Fas-mediated apoptosis than NZB peritoneal B-1 cells. The results presented here raise the possibility that the reduced susceptibility to Fas-triggered apoptosis in B-1 cells might be an accelerating factor for the autoantibody production in NZB mice.     

Human peritoneal B-1 cells and the influence of continuous ambulatory peritoneal dialysis on peritoneal and peripheral blood mononuclear cell (PBMC) composition and immunoglobulin levels

In mice, peritoneal B cells are composed of a unique B-1 cell population which can repopulate the intestinal lamina propria with IgA-producing cells, as well as contribute to the majority of serum IgM. In this study, peritoneal lymphocytes from patients starting continuous ambulatory peritoneal dialysis (CAPD) and from women undergoing bilateral tubal ligation (BTL) were analysed for the presence of a B-1 cell population as well as the expression of potential homing receptors. Up to 63% of the peritoneal B cells express surface antigen CD5, and most peritoneal lymphocytes express the mucosal homing receptors, α4β7 and αEβ7. When analysing serial samples collected from patients from the beginning of dialysis to 1 year, no marked changes were observed in serum or salivary immunoglobulin levels, although the peritoneal lymphocyte population was reduced by 50%. These data suggest that the phenotype of human peritoneal B-1 cells is similar to that of mice, but the contributions to the immune system may differ.     

Mouse B-1 cell-derived mononuclear phagocyte, a novel cellular component of acute non-specific inflammatory exudate

At least three B cell subsets, B-1a, B-1b and B-2, or conventional B cells are present in the mouse periphery. Here we demonstrate that B-1 cells spontaneously proliferate in stationary cultures of normal adherent mouse peritoneal cells. B-1 cells were characterized by morphology, immunohistochemistry and flow cytometry. IgM was detected in the supernatants of these cultures. We demonstrated that the major cell population analyzed expresses the B-1b phenotype. When these cells were transferred to a new culture, a large proportion of them adhere to the plastic surface, and spread as bipolar cells endowed with the capacity to phagocytose via Fc and mannose receptors. Flow cytometry analysis of these adherent cells demonstrated that the great majority of them share both B-220 and Mac-1 antigens. Nevertheless, 45% of them were exclusively Mac-1(+). Finally, when they were labeled in vitro with [(3)H]thymidine and transferred to the peritoneal cavity of naive mice, they migrate to a non-specific inflammatory focus induced by a foreign-body implant. These data demonstrate that B-1 cells, mainly B-1b cells, not only proliferate and differentiate into a mononuclear phagocyte in vitro, but also that they exit the peritoneal cavity and migrate to a non-specific inflammatory milieu.     

Role of B-1a cells in autoimmunity

B-1a cells are distinguished from conventional B cells (B2) by their developmental origin, their surface marker expression and their functions. They were originally identified as a B cell subset of fetal origin that expresses the pan-T cell surface glycoprotein, CD5. B-1a cells also differ from B2 by the expression levels of several surface markers, including IgM, IgD, CD43 and B220 [R. Berland, H.H. Wortis, Origins and functions of B-1 cells with notes on the role of CD5. Ann Rev Immunol, 20 (2002) 253-300.]. The majority of B-1a cells are located in peritoneal and pleural cavities. Compared to B2 cells, B-1a are long-lived, non-circulating, with reduced BCR diversity and affinity [A.B. Kantor, C.E. Merrill, L.A. Herzenberg, J.L. Hillson, An unbiased analysis of V-H-D-J(H) sequences from B-1a, B-1b, and conventional B cells. J Immunol, 158 (1997) 1175-1186.]. B-1a cells are largely responsible for the production of circulating IgM referred to as natural antibodies. These low affinity antibodies are polyreactive and constitute as such a first line of defense against bacterial pathogens [M.C. Carroll, A.P. Prodeus, Linkages of innate and adaptive immunity. Curr Opin Immunol, 10 (1998) 36-40.]. This polyreactivity also results into the recognition of autoantigens, which serves in the clearance of apoptosis products. The weak autoreactivity of the B-1a cells has been postulated to play a role in autoimmune pathogenesis. In addition, other characteristics, such as the production of high level of IL-10 [A. O'Garra, R. Chang, N. Go, R. Hastings, G. Haughton, M. Howard, et al. Ly-1 B (B-1) cells are the main source of B cell-derived interleukin 10. Eur J Immunol, 22 (1992) 711-717.] and enhanced antigen presentation capacities [C. Mohan, L. Morel, P. Yang, E.K. Wakeland, Accumulation of splenic B1a cells with potent antigen-presenting capability in NZM2410 lupus-prone mice. Arthritis and Rheumatism, 41 (1998) 1652-1662.], have implicated B-1a cells in autoimmunity. This review will discuss the current understandings of their role in autoimmune diseases with focus on lupus.     

Unraveling B-1 progenitors

B-1 cells comprise a small percentage of the B lymphocytes that reside in multiple tissues in the mouse, including the peritoneal and pleural cavities. Functionally, B-1 cells participate in innate immunity by producing the majority of the natural IgM in serum, which protects against invading pathogens before the onset of the adaptive immune response. B-1 cells arise from fetal and neonatal progenitors and are distinct from the adult bone marrow progenitors that give rise to follicular and marginal zone B-2 cells. Recent studies have attempted to delineate the progenitors of B-1 cells from those of B-2 cells. Notably, the identification of CD45R(-/lo)CD19(+) B-1 progenitors and expression of two surface determinants, CD138 and major histocompatibility class II antigens, distinguish developing B-1 cells from B-2 cells.     

Morphological characterization of mouse B-1 cells

At least three B cell subsets, B-1a, B-1b and B-2 are present circulating peripherally in the mouse. In these animals, B-1 cells constitute a minor fraction of B cells in spleen and are absent in lymph nodes although they represent the main B cell population in peritoneal and pleural cavities. Currently these cells are identified by a surface phenotypic repertoire; they express Mac-1, IgM(high), and B220(low). B-1a cells express CDS. The aim of this work emerged from the fact that the morphology of B-1 cells is not fully characterized. Here we identified B-1 cells using colloidal gold immunocytochemical assays and purified B-1 cells from supernatants of adherent peritoneal cell cultures by a magnetic bead technique. These techniques lead us to demonstrate that, in mice, either B-1a or B-1b cells have a unique morphology distinct from that of B-2 cells.     

Immunofluorescence analysis of B-1 cell ontogeny in the mouse

In order to further understand the developmental aspects ofB-1 cells, we characterized the ontogeny of this B cell populationin the spleen and peritoneal cavity of BALB/c mice. Althoughthere are B-1 cells in the spleen within the first 1–3weeks after birth, they do not at any stage represent the majorityof splenic B cells. Splenic B-1 cells reach peak levels at 9days after birth. The mesenterlc lining that covers the smallintestine of 7-day-old mice contains a population of IgM⁺ Bcells, while at the same age, there are few lymphold cells inthe peritoneal cavity. Between 7 and 8 days after birth thereis an influx of B cells into the peritoneal cavity. At 8 days,the first detectable peritoneal B cells appear to be of theB-1 type based on expression of IL-5 receptor and CD5. However,these peritoneal B-1 cells do not express Mac-1. This antigenIs not expressed by the majority of peritoneal B-1 calls until3 weeks. This study indicates that the majority of early splenicB cells are not B-1 cells and it suggests that the mesenterlctissues surrounding the gut contain B lymphocytes which trafficinto the peritoneal cavity where they then reside.     

B-1-like cells exist in sheep. Characterization of their phenotype and behaviour.

Two populations of B lymphocytes, B-1 (CD5+ and/or CD11b+) and B-2 (CD5- and CD11b-) cells have been described. In mice, which is the species of reference for B-1 and B-2 cell studies, these two subsets present different developmental schemes, phenotypes, antibody repertoires, localization and behaviours. Interestingly, in sheep, B cells rearrange their immunoglobulin (Ig) loci around the neonatal period, similarly to murine B-1 cells. However, the phenotype of the sheep B cells has not been characterized with regards to their developmental pathway. In this report, we show that two sheep B-cell subsets can be distinguished on the basis of CD11b expression. Relative to CD11b- B cells, the CD11b+ B cells frequently co-express CD5, CD11c, higher levels of surface IgM (sIgM), show larger cell size and higher cell-cycling activity, and thus present a B-1-like phenotype. However, unlike murine B-1 cells, sheep B-1 like cells mainly localize in blood, display a higher propensity to spontaneous apoptosis relative to B-2-like cells, and proliferate after sIgM stimulation. Our data show that despite neonatal immunoglobulin loci rearrangements, sheep B cells do not all express a B-1-like phenotype. However, B-1-and B-2-like cells co-exist and present phenotypic and behavioural specificities. Nevertheless, sheep B-1-and B-2-like cells differ from the murine B-1 and B-2 cells in their cell behaviour. These subsets can thus not be considered as true homologues among species.     

Evidence that intestinal IgA plasma cells in {micro},{varkappa} transgenic mice are derived from B-1 (Ly-1 B) cells

B6-Sp6 transgenic mice carry fully rearranged (BALB/c-derived,Igh-C^a allotype) µ. heavy chain and light chain transgenes,specific for trinitrophenyl, on a C57BL background (Igh-C^b allotype).FACS analyses show that the majority of B cells in peripherallymphoid organs and bone marrow(BM) express transgenic IgM exclusively.A small proportion of the B cells, however, express endogenousIgM, usually concomitant with transgenic IgM. Three criteriaestablish that the endogenous IgM expressing B cells belongto the B-1 cell lineage. (I) Endogenous IgM expressing B cellsin B6-Sp6 mice have the same localization pattern as B-1 cellsfrom normal animals: they are enriched in the peritoneal cavity.(II) The endogenous IgM⁺ B cells have the phenotype of B-1 cells:the endogenous IgM⁺ peritoneal B cells express Mac-1 (CD11b)and low levels of IgD, and most also express CD5 (L-1). (III)B6-Sp6 BM poorly reconstitutes endogenous IgM⁺ B cells, justas adult BM from normal mice poorly reconstitutes B-1 cells.In contrast, B cells which only express the transgene are readilyreconstituted by B6-Sp6 BM. The few endogenous IgM⁺ cells inthe B6-Sp6 BM recipients are located in the peritoneal cavityand have the phenotype of B-1b cells (previously the Ly-1 Bsister population), which are known to be reconstituted by adultBM.Two-color immunofluorescence staining of tissue sectionsfrom the gut and from isolated gut lamina propria cells showsthe presence of many IgA containing cells, about one-third ofwhich simultaneously express cytoplasmic (transgenic) IgM. TheC-region of this IgA is produced by endogenous C a genes, becausethe transgene encodes only for C_µ. Furthermore, the majorityof gut IgA containing cells do not express the Idiotype of thetransgene, indicating that most of the gut IgA cells are encodedby endogenous V_H genes and thus the result of an isotype switchfrom endogenous IgM expressing B cells. Since the endogenousIgM⁺ cells are B-1 cells (both B-1a and B-1b), the data stronglyindicate that the intestinal IgA plasma cells also belong tothe B-1 cell lineage.     

Effects of IgM Allotype Suppression on Serum IgM Levels,B-1 and B-2 Cells,and Antibody Responses ir Allotype Heterozygous F1 Mice

IgM allotype heterozygous F1 mice were independently suppressed for Igh6a or Igh6b to evaluate the contribution of B-1 and B-2 cells to natural serum IgM levels and Ab responses. B-2 B cells expressing IgM of the suppressed allotype were evident in the spleens of suppressed mice 4 to 6 weeks after cessation of the suppression regimen, whereas B-1 B cells of the suppressed allotype were undetectable for up to 9 months. Although serum IgM of the suppressed allotype was initially depleted in mice suppressed for either allotype, by 7 months of age, there were detectable levels of IgM of the suppressed allotype in the serum; however, the levels were significantly below that found in nonsuppressed mice. When mice were immunized with either the T-independent or T-dependent form of phosphorylcholine, those suppressed for either allotype, and consequently depleted of B-1 B cells of that allotype, did not respond with phosphorylcholine-specific IgM of the suppressed allotype. In contrast, when mice were immunized with α1-3 dextran, the Igh6a allotype-suppressed mice were able to produce dextran-specific IgM of that allotype. These results show that allotype-bearing B-1 cells of both allotypes can be effectively suppressed by this suppression protocol and this produces long-lasting effects on B-1 cell levels and serum IgM of the suppressed allotype. These observations reflect the derivation of the majority of B-1 cells from fetal-neonatal precursors, which cannot be replaced by newly emerging B-2 cells of adult origin. Their ablation by antibody treatment results in permanent alterations to the adult B-cell repertoire.     

Expression of mouse CD43 in the B cell lineage of transgenic mice causes impaired immune responses to T-independent antigens

CD43 (leukosialin), a sialylated glycoprotein expressed on the surface of most hematopoietic cells, has been implicated in cell adhesion and signaling. However, its precise physiological function remains unclear. We used mouse CD43 (mCD43)-immunoglobulin enhancer-transgenic (TG) mice to study the role of mCD43 in vivo. Previous work revealed that mCD43 expression on mature B cells in these mice resulted in immunodeficiency to T-dependent (TD) antigens (Ag), possibly by impairing B-T cell interactions. In the present study we have immunized the TG mice with the T-independent (TI) Ag fluorescein-(Fl) lipopolysaccharide (LPS) (TI type 1 Ag) and Fl-Ficoll (TI type 2 Ag). Surprisingly, the mCD43-Ig enhancer expressing mice were impaired in their ability to mount humoral responses to both Fl-LPS and Fl-Ficoll, and had decreased numbers of cells responding to Ag in vivo. Flow cytometric analysis was performed on peritoneal B-1 cells, a population which often plays a major role in humoral responses to TI Ag such as bacterial Ag. This analysis revealed similar B220, IgM and CD5 expression patterns for the TG and nontransgenic (NTG) B-1 cells. In addition, purified peritoneal B-1 cells from TG and NTG mice were able to respond to LPS. Stimulation of splenic B cells in vitro with Fl-LPS and Fl-Ficoll revealed that, in contrast to NTG B cell responses, TG B cell responses could not be enhanced by co-culture with T cells. However, soluble T cell factor enhancement of the TG B cell responses was normal. These data suggest that the mCD43 expression on B cells may inhibit cell interactions that are important for enhanced TI Ag responses. The anti-adhesive forces of mucins in general may thus be critical in regulating both TD and TI humoral responses.     

Acute depletion and recovery of peritoneal B-1 lymphocytes in BALB/c mice after a single injection of mercury chloride

The acute toxicity of mercury (Hg) to B cells was studied in the peritoneal cavity of BALB/c mice, a coelomic space where both B-1 and B-2 subsets of B lymphocytes are present. Up to 24 hr after a single in situ Hg injection, the peritoneal cavity became virtually devoid of lymphocytes, particularly of the B-1 subset. Lymphocyte depletion was more severe for B than T cells. This depletion was associated with partial lymphocyte activation (CD69⁺) at 6 hr of treatment and it was due to apoptosis rather than to necrosis. Partial recovery of both B and T cells was observed in the peritoneal cavity 48 hr after the Hg injection. The phenomenon was followed by a second decrease in peritoneal lymphocytes 72 hr after Hg. Neutrophils that entered the peritoneal cavity because of the Hg injection were resistant to apoptosis. No significant changes in lymphocyte number or subpopulation were found in the spleen and thymus of the mice up to 72 hr after the Hg treatment. We concluded that B lymphocytes were severely affected by the toxic effects of Hg. Our data suggest that Hg-induced unbalance in the repertoire of B cells, of the B-1 subset in particular, may result later in the secretion of the high titres of pathogenic autoantibodies that are found in the Hg-induced lupus disorder of BALB/c mice.